(a) Field of the Invention
The present invention relates to a method for preparing .alpha.-L-aspartyl-L-phenylalanine methyl ester from an acid addition salt thereof.
.alpha.-L-aspartyl-L-phenylalanine methyl ester (hereinafter abbreviated as ".alpha.-APM") is a widely known dipeptide sweetener. It has sweetness of good quality and a high degree of sweetness, i.e., about 200 times the sweetness of sucrose. The demand for .alpha.-APM as a diet sweetener is rapidly expanding.
(b) Description of the Prior Art
.alpha.-APM is a dipeptide composed of L-aspartic acid and L-phenylalanine methyl ester. The known methods for the preparation of .alpha.-APM can be classified into a biochemical process utilizing microorganisms and a chemical process. For each process, various methods have been disclosed.
In the biochemical process, N-benzyloxycarbonyl-L-aspartic acid and L-phenylakanien methyl ester are condensed in the presence of metalloprotease to obtain N-benzyloxycarbonyl-.alpha.-L-aspartyl-L-phenylalanine methyl ester. The benzyloxycarbonyl group is then removed from the intermediate by catalytic reduction to give .alpha.-APM (Japanese Laid-open Patent Publication No. 55-135595).
As a typical example of the chemical process, L-aspartic anhydride having a protected amino group is subjected to a condensation reaction with L-phenylalanine methyl ester in a suitable solvent, and the protective group is then removed therefrom by a conventional method to obtain .alpha.-APM (U.S. Pat. No. 3,786,039).
In another method, N-formyl-L-aspartic anhydride and L-phenylalanine are condensed, and deformylation and esterification are then simultaneously carried out in an acidic medium (Japanese Laid-open Patent Publication No. 53-82752).
When an N-protected-L-aspartic anhydride is used as a starting material, .beta.-L-aspartyl-L-phenylalanine methyl ester (hereinafter abbreviated as ".beta.-APM") is inevitably formed as a by-product in addition to .alpha.-APM. This .beta.-APM has no sweetening effect and instead has a bitter taste, and so it is necessary to remove the .beta.-APM from the thus-produced .alpha.-APM.
In every purification method, removal not only of .beta.-APM but also other impurities from .alpha.-APM is required.
Japanese Patent Publication No. 49-41425 discloses the technique of purifying crude .alpha.-APM containing .beta.-APM as a salt of a hydrohalogenic acid from an aqueous solution.
Furthermore, Japanese Patent Publication No. 51-40071 discloses a process of obtaining high-purity .alpha.-APM which comprises dissolving crude .alpha.-APM containing impurities in an aqueous mineral acid solution, removing 5-benzyl-3,6-dioxo-2-piperazine acetic acid (hereinafter abbreviated as "DKP") which is a reaction by-product, and then effecting neutralization.
In these methods, in order to isolate .alpha.-APM from the reaction solvent or to separate impurities such as .beta.-APM and DKP, a mineral acid salt or an organic sulfonate of .alpha.-APM is formed and then neutralized with a base.
Usually, when isolating .alpha.-APM by neutralizing an acid addition salt of .alpha.-APM, the neutralization is carried out with a neutralizing agent such as an inorganic base, e.g., sodium hydroxide, or an organic base, e.g., triethylamine, in an aqueous solution, and the precipitated .alpha.-APM crystals are then isolated by filtration. As the acid component of a mineral acid salt or the organic sulfonic acid salt of .alpha.-APM, hydrochloric acid, sulfuric acid or p-toluene sulfonic acid is usually used. Therefore, when this salt is neutralized with the inorganic or organic base, at least an equilmolar inorganic or organic salt is present in the mother liquor, and this salt adheres to a filter cake and contaminates the resulting product. The contamination of the product with a salt such as hydrochloride or a sulfate reduces the commercial value of the product. According to the standard specification of usual amino acids, the content of anions such as chlorine ions and sulfate ions should be controlled to a level of 0.02 or 0.03%.
In addition, in this neutralization reaction, dissolved .alpha.-APM is present in a perceptible amount in the filtrate, which is why the yield is low. The filtrate can be reused as the solvent for the neutralization reaction, but in this case, the contained salt of a mineral acid or an organic sulfonic acid therein is accumulates in the solvent, with the result that the amount of the salt in the isolated product increases. Accordingly, a purification operation is necessary to remove the inorganic ions from the filtered and isolated .alpha.-APM.
Since the solubility of .alpha.-APM in a solvent such as water is low, a large amount of the solvent is required when .alpha.-APM is completely dissolved and then treated. In addition, being unstable to heat, .alpha.-APM cannot be treated at a high temperature for a long period of time. Conversely, when .alpha.-APM is treated in a suspension state, the properties of its slurry are so bad that operation such as stirring or liquid transfer is very difficult, and if the stirring is carried out at a high speed, bubbles which are hard to break are formed. Moreover, if a filtration speed is slow, the resulting cake inconveniently contains a large amount of the liquid. Thus, it is industrially very inefficient that the purification operation is repeated in which an .alpha.-APM solution or suspension having the bad operation characteristics is employed.
Furthermore, in the usual neutralization reaction using a soluble base, contact of .alpha.-APM with the base results in the formation of DKP, which deteriorates the yield, and if the product is contaminated with the formed DKP, the commercial value of the product is reduced.
As discussed above, in the conventional neutralization process for the acid salt of .alpha.-APM, the yield is low, and a repurification operation is necessary. Consequently, the above-mentioned process is industrially undesirable.